Grateloupia jejuensis (Halymeniales, Rhodophyta): a new species previously confused with G. elata and G. cornea in Korea
- Author: Kim Su Yeon, Han Eun Gyu, Kim Myung Sook, Park Jung Kwang, Boo Sung Min
- Organization: Kim Su Yeon; Han Eun Gyu; Kim Myung Sook; Park Jung Kwang; Boo Sung Min
- Publish: ALGAE Volume 28, Issue3, p233~240, 15 Sep 2013
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ABSTRACT
Despite specimens’ large size and ease of collection in northeast Asian waters, the species diversity of the genus
Grateloupia still needs more research in Korea. We investigated plastidrbc L sequences and carried out detailed morphological observation on flattened halymeniacean red alga collected in twelve locations around Korea and Japan. We describeGrateloupia jejuensis sp. nov. based on the distinct clade with high support in ourrbc L tree.Grateloupia jejuensis is characterized by solitary or caespitose habit and flattened thalli with discoid holdfast, cartilaginous texture, and blunt or bifid axis.Grateloupia jejuensis was distantly related toG. elata andG. cornea , which have been morphologically confused with the former, and it formed a sister relationship withPrionitis filiformis from California, USA in therbc L tree.
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KEYWORD
Grateloupia , Grateloupia jejuensis , Halymeniales , Korea , new species , Rhodophyta
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The genus
Grateloupia was established by C. Agardh (1822) based onG. filicina (J. V. Lamouroux) C. Agardh.Grateloupia is distinguished byGrateloupia -type auxiliary cell ampullae with a single primary ampullar filament and two or three 7- to 13-celled secondary ampullar filaments of a conical out line (Chiang 1970, Kawaguchi 1989, Wang et al. 2000). Recent studies combining morphological and molecular data have resulted in several taxonomic changes.Pachymeniopsis Y. Yamada,Prionitis J. Agardh,Dermocorynus P. L. Crouan & H. M. Crouan, andPhyllymenia J. Agardh have been merged intoGrateloupia (Kawaguchi 1997, Wang et al. 2001, De Clerck et al. 2005b , Wilkes et al. 2005) whileG. intestinalis (Harvey) Setchell moved toGlaphyrosiphon Hommersand & Leister (Hommersand et al. 2010).Grateloupia elata (Okamura) S. Kawaguchi & H. W. Wang was described based on specimens under the namePrionitis elata Okamura (type locality: Shirahama, Chiba Prefecture, located at the Pacific side of Japan). In Korea, Cotton (1906) reported this species (asPrionitis elata ) for the first time at Wonsan on the northeast coast of the Korean peninsula. Afterwards,G. elata was reported between Busan and Gangneung on the east coast of Korea and Gapado on Jeju Island (Lee and Kang 2001).Grateloupia cornea Okamura (1913) was transferred to the genusCarpopeltis and then changed toPrionitis cornea (Okamura) E. Y. Dawson (Dawson 1958). However, Wang et al. (2001) reinstated the original name,Grateloupia cornea , when they mergedPrionitis intoGrateloupia . In Korea, Kang (1966) reported this species on the east to south coasts and on Jeju Island. Subsequently, this species has been reported in many floristic reports (Lee and Kang 2001).The taxonomy of the genus
Grateloupia remains confused due to the low taxonomic utility of vegetative morphological characteristics, such as surface morphology, thallus habit and texture, and frequency of proliferations (Wang et al. 2001, De Clerk et al. 2005a , Wilkes et al. 2005). Recent studies show that therbc L gene is a suitable marker for taxonomic study ofGrateloupia (Wang et al.2000, Faye et al. 2004, Lee et al. 2009, Yang et al. 2013). In the present study, we analyzed
rbc L from unidentified specimens in Korea as well as other species and observed morphological characters. As a result, we describeGrateloupia jejuensis sp. nov. and discuss the phylogenetic relationships ofG. jejuensis with the other species of the genus.In total, 38 specimens of 13 species in the Halymeniaceae were collected from Korea, China, Japan, Russia, and USA. Field observation and collections of
Grateloupia jejuensis were made at intertidal zone of 12 locations around Korea and Japan from 2009 to 2012.Specimens were sectioned with a freezing microtome (FX-801; Yamato Kohki Industrial Co. Ltd., Tokyo, Japan). Photographs were taken with a DP-71 camera (Olympus, Tokyo, Japan) attached to a microscope (BX 51; Olympus). Voucher specimens are housed at the herbarium of Chungnam National University, Daejeon, Korea (CNUK).
We analyzed 84 sequences representing 48 Halymeniaceae species, including 38 new sequences and 46 GenBank sequences. Information for specimens, their collection data, and GenBank accession numbers of
rbc L sequences are given in Table 1. DNA extraction, PCR amplification, and sequencing follow the procedures in Boo et al. (2010). For amplification and sequencing reaction of therbc L gene, specific primer pairs wererbc LF145-rbc LR898 andrbc LF762-rbc LR1442 (Kim et al. 2010). Electropherogram outputs from each sample were edited using Chromas version 1.45 (http://www.technelysium.com.au/chromas.html). Eighty-ninerbc L sequences were collated and aligned using Se-Al (version 2.0a11), as per Rambaut (2002). Maximum likelihood analyses were conducted using RAxML (Stamatakis 2006) with the GTR + Γ evolutionary model. We performed 200 independent tree inferences using the -# option with default ?I (automatically optimized SPR rearrangement) and ?c (25 distinct rate categories) option in the program to identify the best tree. To generate bootstrap values for the best phylogeny, we used 1,000 replications under the same program with the same model settings.> Grateloupia jejuensis S. Y. Kim, E. G. Han & S. M. Boo sp. nov. (
Fig. 1 )Thallus solitariam aut caespitosa, applanatis et dichotomis calami, saepe flabellatae in adumbration, apice cum terminus in obtunsus aut bifida, cartilaginous in textum. Cortex cum 7-8 compactus corticales cellulis, sphaericus in intimus cellulis et angulata superficies cellulis. Medulla cum parvos sphaericus cellulis, 5-7 μm in diametros. Tetrasporic sporophylls, tenuis et parvos, oblongae, acuminatus ad basin, seriatum per duo marginibus. Tetrasporangi is cruciatis dividuus, plasmatio ex corticales cellam iacuit in the proliferous ramulis. Thallus solitary or caespitose, flattened and dichotomous branches, often flabellate in outline, apex with ending in blunt or bifida, cartilaginous in texture. Cortex with 7-8 compact cortical cells, spherical in innermost cells and angulated surface cells. Medulla with small spherical cells, 5-7 μm in diameter. Tetrasporic sporophylls, thin and small, oblong, tapering at base, seriated along both margins. Tetrasporangia cruciately divided, forming from cortical cell layer in the proliferous branchlets.
Holotype. CNU040218, intertidal zone of Hado (33°31? N 126°54? E), Jeju, Korea, Oct 4, 2009 (Fig. 1A), Herbarium of Chungnam National University (CNUK), Daejeon, Korea. Isotypes: CNU040217, -19, -220-1, NIBRAL0000138259- 60.Etymology. The specific epithet refers to Jeju Island, Korea, where the type was collected.Distribution. Found from Gangneung on the east coast to Jinhae on the south coast, and from Ilkwari to Sungsan around Jeju Island.Grateloupia jejuensis was frequently found throughout the year on rocks in tide pools and sheltered places in lower tidal zones. Specimens were collected in February, June, July, October, and December; tetrasporophytes were collected in May and October. Gametophytes were not found.Korean name. 댓잎도박Morphology. Plants are solitary or caspitose, arising from discoid holdfast, and cartilaginous in texture. The thallus is dark purple to red in color and bright red in the upper portion. The thallus is 8-16 cm high, 0.2-0.4 cm wide, and 200-350 μm thick, with a short stipe (0.5-1 cm long). Branches are divided up to eight times dichotomously, and apex endings are blunt or bifid. The marginal branches produce proliferous branchlets that are often irregular and small (up to 5 mm long and 1 mm wide), scattered over the middle part of the thallus except for the stipe and basal part. The internal thallus is composed of compacted cortex and dense medulla. The cortex is 7-8 cell layers and is 5-20 μm in diameter. The outer cortex is composed of 3-4 layers with narrowly ellipsoidal cells, and the inner cortex is composed of 3-4 layers with irregular or rounded transparent cells. The medulla is composed of filamentous cells (5-7 μm in diameter). Tetrasporangia are formed from the cortical cell layer in the proliferous branchlets. The mature tetrasporangia are cruciately divided, narrowly ellipsoidal in shape, and 30-40 μm long by 10-15 μm wide.Phylogeny of 1,094-nucleotide portions of therbc L.rbc L gene were aligned for 84 sequences (38 new sequences),representing 48 species of Halymeniaceae. All sequences of
G. jejuensis from 12 sites in Korea and Japan were identical. Therbc L difference ofG. jejuensis withPrionitis filiformis Kylin was 14-15 bp (1.28-1.37%), whileG. jejuensis differed by 27-29 bp (2.28-2.45%) fromG. elata and 63 bp (5.48%) fromG. cornea .In the phylogenetic tree (Fig. 2),
Grateloupia jejuensis was placed in a single clade and clearly separated from the other species of the genus. The sister species ofG. jejuensis wasPrionitis filiformis (86% for maximum likelihood).Grateloupia cornea was resolved as a sister taxon ofG. chiangii from Japan with 91% bootstrap support.Grateloupia angusta shows sister relationships withG. elliptica andG. lanceolata .The main finding of the present study is the discovery of a new species,
Grateloupia jejuensis , in Korea based onrbc L sequence data and morphological evidence. It had previously been misidentified asG. elata (Lee 2008) orG. cornea (Lee 1987) because of similarity in cartilaginous texture, and erect, linear, compressed habit. In this context, the existence ofG. elata andG. cornea in Korea may be questionable.The comparative morphology of
Grateloupia jejuensis with similar species is summarized in Table 2.Grateloupia elata is much larger in thallus size thanG. jejuensis (20- 50 cm vs. 8-16 cm). Kawaguchi et al. (2001) reported that the difference in size is the one of main characteristics to distinguishG. asiatica fromG. filicina (10-30 cm vs. 9-12 cm). The width ofG. elata is less than that ofG. jejuensis (1.5-2.5 mm vs. 2-4 mm). The difference in habitat is also a remarkable characteristic between the two species:G. elata occurs in deep waters (Okamura 1899), whereas
G. jejuensis usually occurs in tide pools and / or intertidal zone.Grateloupia cornea is also similar toG. jejuensis in having cartilaginous texture and linear habit. However,G. jejuensis has 7-8 cortical cell layers, whileG. cornea has 13-15 cortical cell layers. The structure of medullary cells ofG. jejuensis is more densely constructed than those ofG. cornea .Okamura (1899) stated that
G. elata resemblesG. angusta (Okamura) S. Kawaguchi & H. W. Wang in structure, habit, and substance, but differed in fruit-bearing (tetraspores and cystocarps) portion and cells of the intermediated layer.G. angusta is distinguished fromG. jejuensis by composition of medulla and number of cortical cells.G. jejuensis has a densely composed medulla with 7-8 cortical cells, whileG. angusta has a very densely composed medulla with 14-16 cortical cells.Wang et al. (2001) merged
Prionitis toGrateloupia ; however, the formal proposals were not for all species ofPrionitis . The genusPrionitis is still used in the Halymeniaceae. In ourrbc L tree,P. filiformis was the sister ofG. jejuensis and grouped together with all otherGrateloupia species with high bootstrap support. It is therefore suggested thatP. filiformis can be transferred toGrateloupia . Before taxonomic revision, a more detailed morphological study ofP. filiformis is needed.In the phylogenetic tree (Fig. 2), ten Korean species were grouped into four different subclades with strong bootstrap support.
Grateloupia jejuensis was included in subclade I.Prionitis filiformis ,P. sternbergii (C. Agardh) J. Agardh,Grateloupia americana from the northeast Pacific andGrateloupia acuminata Holmes,G. asiatica ,G. divaricata ,G. elata ,G. jejuensis ,G. livida (Harvey) Yamada,G. patens (Okamura) S. Kawaguchi & H. W. Wang,G. schmitziana (Okamura) S. Kawaguchi & H. W. Wang fromthe northwest Pacific, were mixed within this subclade, suggesting that many species were likely to have diverged in the north Pacific Ocean.
Grateloupia cornea andG. angusta formed subclade II. Given the morphological similarity toG. jejuensis , the habit similarity is a result of convergence. This result is consistent with previous studies (Wang et al. 2001, De Clerk et al. 2005a , Wilkes et al. 2005, Lee et al. 2009). Thallus habits are homoplasious withinGrateloupia ; thus, additional taxon sampling will provide a better understanding of phylogenetic relationships of the species.-
6. De Clerck O., Gavio B., Fredericq S., Barbara I., Coppejans E. 2005a Systematics of Grateloupia filicina (Halymeniaceae, Rhodophyta), based on rbcL sequence analyses and morphological evidence, including the reinstatement of G. minima and the description of G. capensis sp. nov. [J. Phycol.] Vol.41 P.391-410
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9. Hommersand M. H., Leister G. L., Ramirez M. E., Gabrielson P. W., Nelson W. A. 2010 A morphological and phylogenetic study of Glaphyrosiphon gen. nov. (Halymeniaceae, Rhodophyta) based on Grateloupia intestinalis with descriptions of two new species: Glaphyrosiphon lindaueri from New Zealand and Glaphyrosiphon chilensis from Chile. [Phycologia] Vol.49 P.554-573
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[Table 1.] Materials used in the present study
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[Fig. 1.] Grateloupia jejuensis S. Y. Kim, E. G. Han & S. M. Boo sp. nov. (A) Holotype (CNU040218, tetrasporophyte) of Grateloupia jejuensis collected from Hado, Jeju, Korea on Oct 4, 2009, deposited in the Herbarium of Chungnam National University (CNUK), Daejeon, Korea. (B) Close-up view of tetrasporic sporophylls (ts). (C) Cross section of thallus showing cortex (c) and medullar layers (m). (D) Longitudinal section of thallus showing cortex (c) and medullar layers (m). (E) Cross section of tetrasporic sporophylls. (F) Close-up view of a tetraspore (t). Scale bars represent: A, 2 cm; B, 5 mm; C, 100 μm; D & E, 50 μm; F, 20 μm.
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[Table 2.] Morphological comparison between Grateloupia jejuensis and the morphologically similar species
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[Fig. 2.] Maximum likelihood (ML) tree inferred from rbcL sequences calculated using the GTR + Γ evolution model. Values above branches refer to ML bootstrap values. Bolded samples show the species which was newly analyzed in this study.